Photorespiratory Properties of Mesophyll Protoplasts of Nicotiana plumbaginifolia

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Abstract
The photorespiratory activity of mesophyll protoplasts of Nicotiana plumbaginifolia has been clearly demonstrated by the presence of a Warburg-effect, the occurrence of an important CO2-sensitive O2 uptake and the effect of some photorespiratory inhibitors on photosynthetic activity. At a nonsaturating dissolved inorganic carbon (DIC) concentration (0.1 millimolar), we observed that the rate of CO2 fixation was 60% lower at 50% O2 compared to that measured at 2% O2. Using 18O2 and mass spectrometry, we measured O2 exchange as a function of light intensity and of DIC concentration. Oxygen uptake measured at the CO2 compensation point (47.4 micromoles O2 per hour per milligram chlorophyll) was three-fold higher than that measured at a saturating CO2 concentration. Cyanide or iodoacetamide, inhibitors of the Calvin cycle, were found to reduce the O2 uptake to the same extent as CO2 saturation. We conclude from these results that the major part of the CO2-sensitive O2 uptake is due to photorespiration. Further, we investigated the effect on net photosynthesis of some inhibitors of the glycolate pathway. At CO2 saturation (10 millimolar DIC), 5 millimolar aminoacetonitrile (AAN), and 1 millimolar aminooxyacetate (AOA) did not cause any significant decrease in net photosynthesis. However, when these two inhibitors were added under a period of active photorespiration (10 minutes at the CO2 compensation point at 20% O2), we observed a decrease in the rate of net photosynthesis at 10 millimolar DIC measured afterward (respectively, 18 and 29%). This inhibition did not appear at 2% O2, but was stronger at 50% O2 (40% for AAN and 47% for AOA). With 0.05 millimolar butyl 2-hydroxy-3-butynoate (BHB) or 0.5 millimolar l-methionine-dl-sulfoximine (l-MSO), rates of net photosynthesis at 10 millimolar DIC were decreased by 10 to 15%. Additional decreases were observed after a period at the CO2 compensation point at 20% O2 (30% for BHB and 20% for l-MSO). From the sites of action of the four inhibitors tested, we suggest the inhibition of photosynthesis occurring after a period of active photorespiration to be due to the toxic accumulation of nonmetabolized phosphoglycolate.